Effects Of Aerobic Exercise On Pulmonary Inflammation And Fibrosis Via AMPK/SIRT1/NF-κB Pathway In Doxorubicin-injected Mice

Objective Doxorubicin(Dox),also known as adriamycin,is one of the most widely used anthracycline anticancer drugs.As the number of cancer survivors has increased,the long-term side effects of chemotherapy drugs have become more pronounced,and the dose dependent toxicity of DOX has limited survival.At present,a large number of related data and mechanisms of DOX cardiotoxicity have been studied,but only a few cases of pulmonary toxicity have been reported.Recent studies have shown that chemicals can attenuate DOX-induced lung injury by reducing markers associated with oxidative stress,inflammation and apoptosis.However,it is not clear whether real-time aerobic exercise has a protective effect on the lungs of DOX-treated mice.Therefore,we continue to explore the expression of AMPK/SIRT1/NF-κB signal pathway and proinflammatory factor in the lungs of mice after DOX injection,and then detect the expression of pulmonary fibrosis and remodeling related proteins.Whether the intervention of aerobic exercise,which started at the same time as the DOX and continued after the drug was discontinued,had a protective effect against DOX-induced lung injury.Methods Thirty-six 8-week-old C57BL/6 male mice were randomly assigned to quiet group(SED-NS,N = 6),doxorubicin group(SED-DOX,N = 12),exercise saline group(EXNS,N = 6)and exercise doxorubicin group(EX-DOX,N = 12).Adriamycin was injected once a week for 4 weeks,5 mg/kg,20 mg/kg,and the control mice were injected with the same amount of saline.The aerobic training regimen consisted of 8weeks of moderate intensity treadmill training.The mice in EX-DOX group were injected with doxorubicin and trained at the same time in week 1-4,and only aerobic exercise was performed at week 4-8.After 8 weeks of aerobic exercise,anesthetized mice were taken samples within 48 hours.Lung tissue was examined histologically,including hematoxylin-eosin staining and Masson staining for pulmonary fibrosis,the expression of related proteins in the lung tissue of mice was detected by Western blotting,it includes AMPK/SIRT1/ NF-κB signal pathway-related proteins,TNF-α,IL-6,IL-10,IL-12,Cox-2 and TGF-β,CTGF,CTGF and remodeling proteins.Results(1)Histologic staining showed that the injection of DOX caused a lot of inflammatory infiltration and bleeding points in the lungs of mice,and the deposition area of blue collagen increased obviously,which was consistent with the expression level of related proteins,these results indicate that the model of DOX induced lung injury has been successfully established;(2)Compared with sed-ns group,the expressions of p-ampk α / AMPK α and IL-10 in lung tissue of sed-dox group were significantly decreased(P < 0.01),the expressions of NF-κ B p65 / NF-κ B and MMP-9 were significantly increased(P < 0.01),and the expressions of IL-6,TGF-β and CTGF were significantly increased(P < 0.05);(3)Compared with sed-dox group,the expression of p-ampk α / AMPK α and IL-10 in ex-dox group increased significantly(P < 0.01),the expression of SIRT1 increased significantly(P < 0.05),and the expression of NF-κ B p65 / NF-κ B,TNF-α,IL-6,TGF-β and MMP-9 decreased significantly(P < 0.05);(4)Compared with sed-ns group,the expression of IL-12 and COX-2 in ex-ns group increased significantly(P < 0.01),the expression of p-ampk α / AMPK α and TNF-α increased significantly(P < 0.05),while IL-10 decreased significantly(P< 0.05);(5)Compared with ex-ns group,the expressions of p-ampk α / AMPK α,IL-12 and COX-2 were significantly decreased in ex-dox group(P < 0.01).The expression of AMPK α and NF-κ B in the lung tissue of four groups of mice had no significant change,and the expression of other proteins had no significant difference.Conclusions The results showed that DOX administration inhibited the phosphorylation of AMPK α and the expression of SIRT1 in lung tissue,and increased the acetylation of NF-κB.The expressions of TNF-α,IL-6,IL-12,COX-2 and other inflammatory factors were increased,along with the expressions of TGF-β,CTGF,Mmp-9 and other fibrosis and remodeling proteins.At the same time,the results showed that aerobic exercise decreased the expression of inflammatory factors and fibrosis proteins,which alleviated the adverse effects of DOX administration.Moreover,aerobic exercise activates the phosphorylation of AMPK α and the expression of SIRT1.Therefore,AMPK/SIRT1/NF-κB signaling pathway may be related to the protection of aerobic exercise on DOX.